Contract Services for CFC, Primary Cell, In vitro & In vivo Assays

By: STEMCELL  09-12-2011
Keywords: Assays, Bone Marrow, Primary Cells

Why use primary cells to test your substance of interest?

In vitro assays using primary cells are:
  • quantitative and qualitative
  • robust
  • flexible and customizable
  • clinically relevant, giving more predictive data
  • cost effective (reducing the number of animals required)  
Contract Assay Services can evaluate the effects of your compound on the following cell types and sources:

*Clinical samples from diseased or normal patients can also be evaluated – inquire for more information. FIGURE 1: Illustration of hematopoietic and neural stem and progenitor cells and the mature cell types that they produce.

Hematopoietic in vitro assays

Our most popular service involves the hematopoeitic colony-forming cell (CFC) assay, an assay that has been validated for the determination of maximum tolerated doses (MTD) by ECVAM.


  We use STEMCELL’s MethoCult


product line, recognized as the global gold standard for the detection and quantification of hematopoietic progenitors using CFC assays. 

Hematopoietic CFC assays can be used to examine simultaneously the effects of substances (lead compounds/clinical candidates, mimetics, cytokines or chemokines) on the processes of proliferation and differentiation. 

Hematopoietic progenitors from different species can be assessed:

Evaluation of Stimulation

Substances thought to stimulate and/or enhance proliferation of hematopoietic progenitors can be tested for functionality in the CFC assay.

FIGURE 1: Stimulation by Erythropoietin (EPO)

Stimulation of erythroid progenitor proliferation by erythropoietin (EPO) is both quantifiable (Figure A) and qualitative (Figure B, colony size increases with increasing [EPO]). In vivo models can be used to confirm stimulatory activity of molecules.

Evaluation of Inhibition or Toxicity

Substances thought to inhibit the proliferation and/or differentiation of hematopoietic progenitors can be screened to assess their effects on the different lineages of progenitors: myeloid, erythroid, and/or megakaryocytes using the CFC assay.  Both IC


and IC


values can be generated to rank test compounds.

TABLE: Comparison of myelotoxicity (CFU-GM) in different species FIGURE 2: Determination of IC50 values for 5-Fluorouracil

Dose response curves and IC50 values for both human BM derived erythroid and myeloid progenitors incubated with 5-Fluorouracil FIGURE 3: Human BM Colonies in the presence of a toxic compound

The size of CFCs will change in the presence of an inhibitory compound, serving as a sensitive toxicity assay readout.


Neural in vitro assays

STEMCELL scientists have worked with leaders in the neural stem cell field to develop standardized reagents and assays for the culture of neural stem and progenitor cells. 


 are highly dynamic, three-dimensional floating spheres containing cells from different stages of neural development – neural stem cells, progenitors and mature cells.  These neurospheres are the basis of both our proliferation (NCFC) assay and differentiation assay.

We use the NCFC assay to assess the effects of your substance of interest on primary neural cells.

FIGURE 1: Assays for neural stem and progenitor cells



Mesenchymal in vitro assays

Drug candidates destined for the tissue engineering market can be assessed for stimulatory or inhibitory effects on the bone marrow microenvironment (mesenchymal cell matrix). These cells, under the appropriate conditions, can differentiate into structural cell types that make up adipose, cartilage, bone and muscle. 

Functional assays using mesenchymal stem and progenitor cells from human bone marrow or mouse compact bone can be used to quantify and qualitatively assess the effects of compounds:

  • Measure the effects of test compound on progenitor frequency
  • Assess progenitor proliferative potential or overall expansion potential
  • Quantitation of mesenchymal progenitor content in bone marrow
  • Test the differentiation potential of mesenchymal stem cells into adipose, osteogenic and chrondrocyte lineages

The Contract Assay Services group has developed new assays for the quantification of mouse CFU-F from compact bone, a source known to be richer than the bone marrow in mesenchymal stem and progenitor cells. Mouse CFU-F frequency increases when culturing compact bone-derived MSCs under low oxygen conditions.

FIGURE 1: Effects of Doxorubicin on Human CFU-F

Dose response curve and IC50 value for human BM derived CFU-F incubated with Doxorubicin. Inhibitory compounds can affect the size and density of the CFU-F colony.

Mobilization, Engraftment and Hematopoietic Cell Recovery

Leverage our

in vivo

expertise to examine the effects of your substance of interest. Our scientists have experience and expertise in the analysis of hematopoietic cell populations in a number of

in vivo


Experimental designs can address mobilization of hematopoietic stem cells from BM into PB, increases in total blood cell counts, and amplification of progenitors and stem cells. 

We perform studies to examine the effects of mobilizing compounds (e.g. G-SCF and AMD3100) as described by Broxmeyer

et al

. The kinetics of tri-lineage (myeloid, erythroid and lymphoid) engraftment in an animal model can also be followed after induction of myelosuppression.  Studies can be customized to evaluate changes in HSC frequency.

TABLE 1:  Mobilization of progenitors and increased erythropoiesis induced by EPO

Wild type BDF1 mice were treated with EPO on days 0 and 1. At day 4 the mice were sacrificed and the progenitor content of the spleens was assessed.  *p<0.005
FIGURE 1: Recovery of CFC after injection of 5-Fluorouracil with and without G-CSF pre-treatment

Wild type BalbC mice were treated with 5-Fluorouracil on day 0. Animals treated with G-CSF (orange line) showed faster recovery of neutrophils in the blood (upper figure) and total progenitors (lower figure) in the bone marrow.


Phenotypic characterization can be performed on normal, transgenic and knockout mice.  Typical analysis includes blood and femoral cell counts, progenitor proliferative studies, stress/recovery experimentation and immunophenotypic analysis.

Keywords: Assays, Bone Marrow, Primary Cells

Other products and services from STEMCELL


Stem Cell Assay Contract Services, Training & Education

STEMCELL Technologies provides a wide variety of training and educational activities to help with the standardization of various stem cell culture assays. Our hands-on training courses provide extensive instruction in the culture of many different cell types and are taught by experts in their field.